LOCUS       sequence-222844-        5977 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of Htt fragment containing amino acids 2-90, 
            with A2C cysteine mutation and a 49 polyQ repeat.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5977)
  AUTHORS   Warner JB 4th, Ruff KM, Tan PS, Lemke EA, Pappu RV, Lashuel HA
  TITLE     Monomeric Huntingtin Exon 1 Has Similar Overall Structural Features 
            for Wild-Type and Pathological Polyglutamine Lengths.
  JOURNAL   J Am Chem Soc. 2017 Oct 18;139(41):14456-14469. doi: 
            10.1021/jacs.7b06659. Epub 2017 Oct 9.
  PUBMED    28937758
REFERENCE   2  (bases 1 to 5977)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1021/jacs.7b06659"; journalName: "J Am Chem Soc"; date: 
            "2017-10-18- 18"; volume: "139"; issue: "41"; pages: "14456-14469"
COMMENT     Sequence Label: pTWIN1-His6-Ssp-Htt2-90(A2C)-49Q
FEATURES             Location/Qualifiers
     source          1..5977
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(810..828)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      824..871
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        931..1035
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1036..1104,1105..1896)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1254..1273)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(1938..2451)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2133..2154)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2345..2364
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      2562..3150
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3051..3070
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3304..3321
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(3343..3362)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3503..3525
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3519..3710)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4267..5349)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     5311..5330
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5350..5427)
                     /gene="lacI"
                     /label=lacI promoter
     terminator      5580..5666
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     5801..5820
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5801..5819
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5820..5844
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     RBS             5859..5881
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             5892..5909
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
ORIGIN
        1 tctgctggat gagaaagatt ttgagatttg ggccattaac gagcagacca tgaaactgga
       61 aagcgcaaaa gttagccgtg ttttttgcac cggtaaaaaa ctggtgtata tcctgaaaac
      121 ccgtctgggt cgtaccatta aagcaaccgc aaatcatcgt tttctgacca ttgatggttg
      181 gaaacgtctg gatgaactga gcctgaaaga acatattgca ctgcctcgta aactggaaag
      241 tagcagcctg caactgagtc cggaaattga aaaactgagc cagagcgata tctattggga
      301 tagcattgtt agcattaccg aaaccggtgt tgaagaagtt tttgatctga ccgttccggg
      361 tccgcacaat tttgttgcaa atgatattat tgtgcacaac tgcaccctgg aaaaactgat
      421 gaaagcattt gaaagcctga aaagctttca gcagcagcaa caacagcagc aacagcaaca
      481 gcagcagcag cagcaacaac agcagcagca acagcaacaa caacaacaac agcagcaaca
      541 acagcaacag cagcaacaac aacagcaaca gcagcagcag caacaacagc aacagcctcc
      601 gcctccgcca ccgcctcctc cacctccgca gctgccgcaa cctcctcctc aggcacagcc
      661 gctgctgccg cagcctcaac ctcctccacc acctccacct ccaccgcctg gtccggcagt
      721 tgcagaagaa ccgctgcatc gtccgtaact gcaggaaggg gatccggctg ctaacaaagc
      781 ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg
      841 ggcctctaaa cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggataact
      901 acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
      961 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     1021 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     1081 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     1141 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     1201 gagagttttc gccccgaaga acgtttccca atgatgagca cttttaaagt tctgctatgt
     1261 ggcgcggtat tatcccgtgt tgacgccggg caagagcaac tcggtcgccg catacactat
     1321 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     1381 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     1441 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     1501 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     1561 cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa
     1621 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     1681 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     1741 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     1801 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     1861 gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat
     1921 atactttaga ttgatttacc ccggttgata atcagaaaag ccccaaaaac aggaagattg
     1981 tataagcaaa tatttaaatt gtaaacgtta atattttgtt aaaattcgcg ttaaattttt
     2041 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     2101 aagaatagcc cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     2161 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     2221 gtgaaccatc acccaaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     2281 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     2341 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     2401 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg taaaaggatc
     2461 taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc
     2521 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     2581 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     2641 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     2701 aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     2761 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     2821 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     2881 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     2941 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     3001 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     3061 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     3121 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     3181 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     3241 gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag
     3301 cgcagcgagt cagtgagcga ggaagctatg gtgcactctc agtacaatct gctctgatgc
     3361 cgcatagtta agccagtata cactccgcta tcgctacgtg actgggtcat ggctgcgccc
     3421 cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct
     3481 tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca
     3541 ccgaaacgcg cgaggcagct gcggtaaagc tcatcagcgt ggtcgtgcag cgattcacag
     3601 atgtctgcct gttcatccgc gtccagctcg ttgagtttct ccagaagcgt taatgtctgg
     3661 cttctgataa agcgggccat gttaagggcg gttttttcct gtttggtcac tgatgcctcc
     3721 gtgtaagggg gatttctgtt catgggggta atgataccga tgaaacgaga gaggatgctc
     3781 acgatacggg ttactgatga tgaacatgcc cggttactgg aacgttgtga gggtaaacaa
     3841 ctggcggtat ggatgcggcg ggaccagaga aaaatcactc agggtcaatg ccagccgaac
     3901 gccagcaaga cgtagcccag cgcgtcggcc gccatgccgg cgataatggc ctgcttctcg
     3961 ccgaaacgtt tggtggcggg accagtgacg aaggcttgag cgagggcgtg caagattccg
     4021 aataccgcaa gcgacaggcc gatcatcgtc gcgctccagc gaaagcggtc ctcgccgaaa
     4081 atgacccaga gcgctgccgg cacctgtcct acgagttgca tgataaagaa gacagtcata
     4141 agtgcggcga cgatagtcat gccccgcgcc caccggaagg agctgactgg gttgaaggct
     4201 ctcaagggca tcggtcgaga tcccggtgcc taatgagtga gctaacttac attaattgcg
     4261 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     4321 ggccaacgcg cggggagagg cggtttgcgt attgggcgcc agggtggttt ttcttttcac
     4381 cagtgagacg ggcaacagct gattgccctt caccgcctgg ccctgagaga gttgcagcaa
     4441 gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg
     4501 gatataacat gagctgtctt cggtatcgtc gtatcccact accgagatat ccgcaccaac
     4561 gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc gccatctgat cgttggcaac
     4621 cagcatcgca gtgggaacga tgccctcatt cagcatttgc atggtttgtt gaaaaccgga
     4681 catggcactc cagtcgcctt cccgttccgc tatcggctga atttgattgc gagtgagata
     4741 tttatgccag ccagccagac gcagacgcgc cgagacagaa cttaatgggc ccgctaacag
     4801 cgcgatttgc tggtgaccca atgcgaccag atgctccacg cccagtcgcg taccgtcttc
     4861 atgggagaaa ataatactgt tgatgggtgt ctggtcagag acatcaagaa ataacgccgg
     4921 aacattagtg caggcagctt ccacagcaat ggcatcctgg tcatccagcg gatagttaat
     4981 gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc gccgctttac aggcttcgac
     5041 gccgcttcgt tctaccatcg acaccaccac gctggcaccc agttgatcgg cgcgagattt
     5101 aatcgccgcg acaatttgcg acggcgcgtg cagggccaga ctggaggtgg caacgccaat
     5161 cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg ttgggaatgt aattcagctc
     5221 cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa acgtggctgg cctggttcac
     5281 cacgcgggaa acggtctgat aagagacacc ggcatactct gcgacatcgt ataacgttac
     5341 tggtttcaca ttcaccaccc tgaattgact ctcttccggg cgctatcatg ccataccgcg
     5401 aaaggttttg cgccattcga tggtgtccgg gatctcgacg ctctccctta tgcgactcct
     5461 gcattaggaa gcagcccagt agtaggttga ggccgttgag caccgccgcc gcaaggaatg
     5521 gtgcatgccg gcatgccgcc ctttcgtctt caagaattaa ttcccaattc cccaggcatc
     5581 aaataaaacg aaaggctcag tcgaaagact gggcctttcg ttttatctgt tgtttgtcgg
     5641 tgaacgctct cctgagtagg acaaatccgc cgggagcgga tttgaacgtt gcgaagcaac
     5701 ggcccggagg gtggcgggca ggacgcccgc cataaactgc caggaattgg ggatcggaat
     5761 taattcccgg tttaaaccgg ggatctcgat cccgcgaaat taatacgact cactataggg
     5821 gaattgtgag cggataacaa ttcccctcta gaaataattt tgtttaactt taagaaggag
     5881 atatacatat gcatcatcat catcaccata gcggtgcaat tagcggtgat agcctgatta
     5941 gcctggcaag caccggtaaa cgtgttagca ttaaaga
//
